Newly synthesized proteins must fold into the correct 3-dimensional conformation in order to function properly in the cell; however, protein folding is a complex and error-prone process. Terminally misfolded proteins that accumulate in the cell or organism can have toxic effects, and therefore must be recognized and promptly removed. ER-associated degradation (ERAD), a protein quality control system that is essential for cellular homeostasis, identifies and degrades terminally misfolded proteins in the endoplasmic reticulum (ER). Defects in ERAD have been linked to numerous human diseases including cystic fibrosis and neurodegenerative disorders; thus, understanding the molecular details of this system may illuminate the underlying pathologies of multiple diseases and reveal new therapeutic targets for the treatment of these diseases. Previous studies of mammalian ERAD have identified some of the components of this system and suggest that ERAD functions as a dynamic network of physically and functionally connected protein complexes. The long-term goal of this study is to gain detailed insight into the organization of the metazoan ERAD network, and understand how this organizational structure allows the system to monitor the folding status of a highly diverse mammalian proteome. The immediate goal of this proposal is to use genetic interaction (GI) mapping to perform a systems-level analysis of the ERAD network topology. The focus of Specific Aim 1 is to perform a genome-wide RNAi screen to identify genes involved in ERAD substrate dislocation. In Specific Aim 2, GIs in the ERAD system will be measured and organized by hierarchical clustering to define network organization and relationships between genes. In Specific Aim 3, the dynamics of the ERAD network will be studied using functional genomics and differential GI analysis. Together, these studies will be used to build a dynamic network map of the mammalian ERAD system.